Chemotherapy is a mainstay for treatment of many human tumors, but in vivo efficacy against quiescent or slowly dividing cancers is poor (Takimoto et al. Cancer Management Handbook (11th Edition), UBM Medica 2009; Sang et al. Trends Mol Med 2010; 16(1):17-26; Mellor et al. Br J Cancer 2005; 93(3):302-9). Radiotherapy and systemically administered chemotherapy achieve specificity by disrupting DNA replication, but cannot ablate quiescent tumor tissues that cycle intermittently. The inability to destroy nondividing tumor cells (including a putative tumor stem cell compartment) is acknowledged as one reason for failure against common human malignancies, including low-growth fraction tumors of prostate, breast, lung, and colon, among many others (Vessella et al. Cancer Biol Ther 2007; 6(9):1496-504; Kusumbe et al. Cancer Res 2009; 69(24):9245-53). Even in the case of a solid tumor with an uncharacteristically high mitotic index (e.g. growth fraction ˜40%), and assuming that all dividing cancer cells are completely destroyed by a cumulative exposure to conventional chemo- or radiotherapy, the mass would still be less than one doubling away from achieving pretreatment dimensions.
Non-metastatic cancers of breast, prostate, larynx, and brain are commonly treated with preoperative radiation therapy (XRT) as a debulking measure prior to definitive surgical resection (DeVita et al. Principles and Practice of Oncology (8th Edition). Ronald A. DePinho and Robert A. Weinbert, Eds. Lippincott Williams & Wilkins, 2008). Other locally invasive, non-metastatic tumors are suitable for life-prolonging XRT, and inoperable malignancies that obstruct a viscus (e.g., stomach, larynx, colon, or airway) are routinely treated with local radiotherapy for palliation (Washington et al. Principles and Practice of Radiation Therapy (3rd Edition). Mosby, 2009). Tumors such as these invariably exhibit a low growth fraction, and at some point become unresponsive to both radio- and the best available chemotherapies.
Several newer modalities have been advanced in an attempt to improve treatment of locally invasive, non-metastatic tumors, including common cancers such as those described above. Cryogenic, magnetic, thermal, and ultrasonic cell ablative technologies, for example, have all been investigated with varying degrees of preclinical or early clinical success (Osada et al. Anticancer Res 2009; 29(12):5203-9; Krishnan et al. Int J Hyperthermia 2010 Sep. 21 [Epub ahead of print]; Margreiter et al. J Endourol 2010; 24(5): 745-6). Experimental gene therapies, such as GDEPT (gene directed enzyme prodrug therapy; so-called “suicide gene” strategies), have been extensively tested, but have met with limited success against locally invasive, non-metastatic tumors for at least two reasons (G W Both. Curr Opin Mol Ther 2009; 11(4): 421-32; Altaner et al. Cancer Lett 2008; 270(2): 191-201; Dachs et al. Molecules 2009; 14(11): 4517-45). First, the efficiency of tumor cell transduction is low with currently available gene transfer vectors. The small proportion of malignant cells that express an anticancer transgene is often not adequate to mount a robust bystander effect against untransduced cells in the tumor mass. Second, GDEPT has primarily utilized the herpes simplex virus thymidine kinase (HSV-tk) gene or prokaryotic cytosine deaminase (CD) gene to activate intratumoral chemotherapy, and the compounds produced by these two enzymes (gancyclovir monophosphate and 5-fluorouracil (FUra), respectively) are primarily active against dividing tumor cells. Low transduction efficiency, poor bystander activity, and failure to kill nondividing cancer cells account for the failure of first generation GDEPT approaches against non-metastatic, solid tumors in the clinic.
The E. coli purine nucleoside phosphorylase (PNP) gene has been shown to generate highly potent compounds such as 2-fluoroadenine (F-Ade) or 6-methylpurine (MeP) intratumorally (Ungerechts et al. Cancer Res. 2007; 67: 10939-10947; Fu et al. Cancer Gene Ther. 2008; 15: 474-484; Fu W, Lan et al. Cancer Sci. 2008; 99: 1172-1179; Parker et al. Cancer Gene Therapy 2011 June; 18(6):390-8; Gadi et al. J. Pharmacol. Exp. Ther. 2003; 304: 1280-1284). Purine bases such as these diffuse freely between E. coli PNP transduced and neighboring (bystander) cells via facilitated diffusion pathways ubiquitous in all cells, and confer a pronounced bystander killing effect (Hong et al. Cancer Res. 2004; 64: 6610-6615). The compounds act by a unique mechanism that disrupts RNA and protein synthesis, and are therefore active against both dividing and nondividing (quiescent) tumor cells in vivo (Parker et al. Biochem. Pharmacol. 1998; 55: 1673-1681). F-Ade can be generated by intracellular E. coli PNP from prodrugs such as 2-F-2′-deoxyadenosine (F-dAdo) or fludarabine phosphate (F-araAMP) (Hong et al. Cancer Res. 2004; 64: 6610-6615; Parker et al. Biochem. Pharmacol. 1998; 55: 1673-1681; Martiniello-Wilks et al. Human Gene Therapy 1998; 9: 1617-1626; Mohr et al. Hepatology 2000; 31: 606-614; Voeks et al. Gene Therapy 2002; 9: 759-768; Martiniello-Wilks et al. J. Gene Med. 2004; 6: 1343-1357; Parker et al. Cancer Gene Therapy 2003; 10: 23-29; Parker et al. Human Gene Therapy 1997; 8: 1637-1644; Martiniello-Wilks et al. J. Gene Med. 2004; 6: 43-54). The latter agent, F-araAMP, is clinically approved for treatment of chronic lymphocytic leukemia, but has no activity against non-lymphoid malignancies.
F-Ade is approximately 1,000 times more active as an anticancer agent than FUra. Despite this potency, numerous laboratories have shown that F-Ade can be used safely as part of GDEPT. Because of 1) strong intratumoral sequestration into cellular nucleic acid, 2) slow release into the systemic compartment following tumor cell death, with uptake by neighboring (bystander) cancer cells in the immediate vicinity, and 3) extensive dilution (throughout the host) of any chemotherapy released from the tumor, the approach leads to safe and consistent antitumor efficacy in numerous animal models in vivo (Ungerechts et al. Cancer Res. 2007; 67: 10939-10947; Fu et al. Cancer Gene Ther. 2008; 15: 474-484; Parker et al. Cancer Gene Therapy 2011 June; 18(6):390-8; Gadi et al. J. Pharmacol. Exp. Ther. 2003; 304: 1280-1284; Hong et al. Cancer Res. 2004; 64: 6610-6615; Martiniello-Wilks et al. Human Gene Therapy 1998; 9: 1617-1626; Mohr et al. Hepatology 2000; 31: 606-614; Voeks et al. Gene Therapy 2002; 9: 759-768; Martiniello-Wilks et al. J. Gene Med. 2004; 6: 1343-1357; Parker et al. Cancer Gene Therapy 2003; 10: 23-29; Parker et al. Human Gene Therapy 1997; 8: 1637-1644; Martiniello-Wilks et al. J. Gene Med. 2004; 6: 43-54; Deharvengt et al. Int. J. Oncol. 2007; 30: 1397-1406; Kikuchi et al. Cancer Gene Ther. 2007; 14: 279-86). Several direct comparisons between E. coli PNP and first generation strategies (HSV-tk and CD) indicate substantial augmentation of GDEPT by a PNP based mechanism (Martiniello-Wilks et al. Human Gene Therapy 1998; 9: 1617-1626; et al. Clin Cancer Res 1997; 3:2075-80; Nestler et al. Gene Therapy 1997; 4:1270-77; Puhlmann et al. Human Gene Therapy 1999; 10: 649-657). The approach has recently been approved by the Food and Drug Administration for clinical testing in the United States (IND #14271, approved Mar. 19, 2010).
The prolonged intratumoral half-life of F-Ade metabolites specifically following generation by E. coli PNP (>24 hours). (Hong et al. Cancer Res. 2004; 64: 6610-6615; Parker et al. Biochem. Pharmacol. 1998; 55: 1673-1681; Parker et al. Cancer Gene Therapy 2003; 10: 23-29) together with bystander killing of quiescent tumor cells and tumor stem cells (by ablating RNA and protein synthesis), suggested the possible use of PNP as a “point and ablate” modality for concentrating potent chemotherapy within tumor tissues.
Cancer treatments with chemotherapeutic drugs have relied on systemic administration as being equivalent or superior to intratumoral injection. Direct intratumoral injection of a chemotherapeutic is not considered by the conventional prior art as being anymore efficacious than systemic routes in eliciting an antitumor effect because of poor intratumoral uptake, poor tumor cell utilization of the chemotherapeutic, and negligible tumor cell lethality in vivo. Additionally, intratumoral administration is an exacting procedure while systemic administration via intravenous or other systemic route is comparatively simple to perform.
Thus, there exists a need to provide a more effective inhibition therapy against in vivo target cells and in particular tumor cells. There further exists a need to improve a bystander inhibitory effect against target cells and to maintain such effect for a prolonged period.